Quantitative analysis of a biological sample of unknown quantity

ABSTRACT

Disclosed is a method for testing a modified specimen such as a dried blood spot, plasma or serum specimen, for an analyte of interest, such as cholesterol. In accordance with the disclosed subject matter, the level of the analyte of interest in the medium from which the modified specimen was obtained (e.g., from a patient&#39;s blood) is determined based on the level of an analyte in a solution formed from the modified specimen and on the level of at least one normalizing analyte. The analyte and normalizing analyte each may be an ion, compound, biochemical entity, or property of the specimen. Also disclosed are a fluid collector and a fluid collection device.

RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.12/343,847 filed on Dec. 24, 2008, which is a divisional of U.S.application Ser. No. 10/421,086, filed on Apr. 23, 2003, which claimspriority to prior application Ser. No. 60/374,629 filed Apr. 23, 2002.The prior application is hereby incorporated by reference in itsentirety.

TECHNICAL FIELD OF THE INVENTION

The invention is in the field of testing, in particular quantitativetesting, and in preferred embodiments medical testing. In highlypreferred embodiments, the invention is directed towards the testing ofbody fluid specimens, in particular blood or serum specimens.

BACKGROUND OF THE INVENTION

Modern medical and wellness practices increasingly make use ofself-administered tests and self-collection of test specimens. Forinstance, U.S. Pat. Nos. 5,978,466; 6,014,438; 6,016,345; and 6,226,378,issued to Richard Quattrocchi and assigned to Home Access HealthCorporation of Hoffman Estates, Ill., all disclose a method ofanonymously testing for a human malady. In accordance with certainembodiments of the subject matter disclosed in the foregoing patents, apatient obtains a blood specimen, typically by pricking his or herfinger, and allows the blood to wick onto a blood spot card. After thecard has dried, the user then sends the blood spot card to a medicaltesting facility, where it is tested to determine whether the patient isafflicted with a specific malady. The user may contact the facilityanonymously to receive the test result.

The subject matter of the foregoing patents is usable in connection withtesting for the presence of human antibodies directed against viralantigens in the blood, for instance, in determining whether a patient isinfected with HIV (human immuno-deficiency virus) or with a hepatitisvirus. Another document, U.S. Pat. No. 5,435,970, issued to Mamenta etal. and assigned to Environmental Diagnostics, Inc. of Burlington, N.C.,discloses a device for separating blood cells from biological fluids,for instance, for separating serum from whole blood. The devicedisclosed in the '970 patent purports to enable the shipment and testingof a serum sample.

The blood spot and serum specimen cards known in the art are suitablefor use in the collection of specimens for qualitative testing, i.e.,testing for the presence or absence of a given compound in blood or agiven medical condition. Heretofore, however, such blood spot and serumcards have been somewhat unsatisfactory in the quantitative testing ofblood and serum specimens.

For instance, general wellness protocol indicates the measurements of apatient's total cholesterol value, which is the number of milligrams oftotal cholesterol in a deciliter of blood. The value is often used inconjunction with a full lipid profile, which provides levels oftriglycerides, HDL (high density lipoprotein) cholesterol, and LDL (lowdensity lipoprotein) cholesterol in a patient's blood. It can be verydifficult to gauge the amount of blood or serum that is present in theblood or serum spot card. Particularly when the blood or serum spot cardhas been self-prepared by a person without medical training, it isdifficult to know to certainty whether the spot card has been“underfilled” with less than the intended quantity of blood or serum or“overfilled” with more than the intended quantity. If the amount ofblood and serum varies by even a small amount over or under the expectedlevel, the usefulness of the quantitative test can be severelydiminished. For instance, it is generally thought that a person's totalcholesterol number should be under 200 mg/dl, with cholesterol numbersabove 240 mg/dl being considered high and with intermediate cholesterolnumber being deemed borderline. A 10% margin of error in a cholesteroldetermination of 220 mg/dl provides no information as to whether theperson's cholesterol level is low, intermediate, or high.

In recognition of these problems, the prior art has provided attempts toprovide a quantitative determination of analyte levels in a bloodspecimen. For instance, U.S. Pat. No. 6,040,135, issued to Steven Tyrelland assigned to Biosafe Laboratories, Inc., Chicago, Ill., purports todisclose a method for correcting for blood volume in a serum analytedetermination. The method that is purportedly disclosed by this documentis limited and is believed generally to be somewhat unsatisfactory.

The invention seeks to improve upon prior art testing methods, and toprovide a method for quantitative testing of modified specimens such asdried blood spot and dried serum specimens.

THE INVENTION

The invention provides multiple embodiments in the field of testing, inparticular medical testing. In accordance with the invention, a modifiedspecimen, preferably a dried blood fluid sample, such as a dried serumor dried whole blood specimen of unknown quantity, is eluted(re-solubilized) and then tested for an analyte. The level of analyte inthe blood from which the modified blood specimen was obtained isdetermined from the level of analyte in a solution formed from the bloodspecimen. A normalizing analyte, which in the preferred embodiment issodium ion, chloride ion, and/or osmolality, is measured and is used inconjunction with the solution level of analyte to determine the level ofanalyte in the blood from which the modified specimen was obtained. Theinvention is not limited to the field of medical testing but, to thecontrary, is useful in connection with other forms of testing. Theinvention further provides methods for preparing a database of testresults, for preparing a regression using a database of test results,and for providing test results to a user.

In alternative embodiments the invention further encompasses a fluidcollector that includes an absorbent substrate coated with a saccharide.A device that includes the collector (as described hereinbelow) also isencompassed by these embodiments.

Other features of preferred embodiments of the invention are set forthhereinbelow.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 is a flowchart representing steps in a method for calculating thelevel of an analyte in blood from which a blood specimen was obtained.

FIG. 2 is a flowchart representing steps in an alternative method forcalculating the level of an analyte in blood from which a blood specimenwas obtained.

FIG. 3 is a flowchart representing steps in a method for providing testresult information to a user.

FIG. 4 is a representation of a database record correlating test resultinformation with a test number.

FIG. 5 is a flowchart representing steps in a method for preparing adatabase of test results and test numbers.

FIG. 6 is a flowchart representing steps in a method for preparing adatabase of blood analyte levels, solution analyte levels, and solutionnormalizing analyte levels.

FIG. 7 is a representation of a database record for a databasecontaining blood analyte level information, solution analyte levelinformation and solution normalizing analyte level information.

FIG. 8 is a schematic illustration showing various communicationsbetween a customer, a results providing facility, and others inconnection with a testing protocol.

FIG. 9 is a perspective view of the obverse side of a blood collectiondevice useful in conjunction with the invention.

FIG. 10 is a perspective view of the reverse side of the device shown inFIG. 9.

DESCRIPTION OF THE PREFERRED EMBODIMENTS

The invention is applicable to the testing of any specimen that ismodified from its original form prior to testing. Most commonly, thespecimen is a dried specimen, which has been dried to facilitate storageor transport of the specimen or for other purposes. In preferredembodiments of the invention, the specimen is a medical specimen, and inhighly preferred embodiments of the invention, the specimen is a bloodfluid specimen, by which is contemplated a dried blood spot, a driedserum spot (for instance, as obtained from the device disclosed in U.S.Pat. No. 5,435,970 or that shown in U.S. Pat. No. 4,839,296 issued toKennedy et al. and assigned to Chem-Elec, Inc. of North Webster, Ind.),or another blood fluid specimen. The invention is applicable to thetesting of the modified specimen for any suitable purpose, and inparticular to testing for any analyte in the specimen. For instance,when the specimen is a blood fluid specimen, the test may be a test forprostate specific antigen (PSA), alanineamino transferase (ALT), lipids,such as triglycerides, high density lipoprotein (HDL), low densitylipoprotein (LDL), or any other analyte of interest. The invention isapplicable to the determination of the level of analyte in the originalspecimen, for instance, the level of total cholesterol in the blood fromwhich a blood fluid specimen has been obtained. The “level” of theanalyte can be expressed in any suitable units, such as molarconcentration, weight concentration, or the like. Blood serum isparticularly preferred, but it is contemplated that other fractions suchas cells, platelets, gamma globulins, plasma or the like may beemployed. More generally, any body fluid is susceptible to analysis inconjunction with the invention. In light of the foregoing, the preferredembodiments of the invention will be further described with respect tothe determination of the lipid profile in a blood sample, but it shouldbe understood that the invention is not limited thereto.

The facility or other entity that performs the test of the blood fluidspecimen may or may not be the same entity that calculates the level ofthe analyte in the blood fluid specimen or the entity that receives aninquiry from a user and reports the test results to the user. To testthe blood fluid specimen, the specimen is first received by the testingentity and is eluted with a liquid, preferably deionized water. It iscontemplated that the liquid may be a non-aqueous liquid or may be anaqueous solution, preferably a solution that is free or essentially freeof sodium ions or any other normalizing analyte. Alternatively, thesolution may have a known amount of the normalizing analyte that can betaken into account during normalization. Preferably, when the testingentity is a testing facility that is intended to test numerousspecimens, the eluant is added in a standard amount, which typically is600 μl (0.6 ml). The eluant in some embodiments may be a bufferedelectrolyte solution.

After eluting the specimen, preferably the specimen first is tested forthe content of a normalizing analyte, such as sodium and chloridecontent, and in some embodiments osmolality, which generally representstotal content of sodium, glucose, and blood urea nitrogen (BUN). To testfor sodium and chloride, an ion specific electrode (ISE), such as thatsold by Orion may be employed. Preferably, information concerning boththe sodium and the chloride content of the solution are obtained, theinformation being, for instance, analog information such as anelectrical signal or digital information such as a printout representingthe sodium or chloride content or a digital signal containinginformation concerning the sodium or chloride content. Most preferably,osmolality also is measured. It should be noted that the invention isnot limited to the use of sodium or chloride as normalizing analytes,but to the contrary, any other analyte (which includes a property suchas osmolality) may be measured. It is contemplated in preferredembodiments that the sodium, chloride, and osmolality levels aremeasured against a predetermined range to determine whether the amountof serum is sufficient to perform an adequate test. For instance, it iscontemplated that for a cholesterol test, there ideally should be aleast approximately 15-17 μl of serum available for testing. If thesodium content of the eluted solution demonstrates that the serum levelis far outside this range, the specimen may be rejected as unsuitablefor testing. Generally, the specimen may be rejected if there isinsufficient serum in the solution, although it is contemplated that insome cases excess serum may be grounds for rejection. Persons skilled inthe art may determine how far outside of the desired range the contentof normalizing analyte may be allowed to vary without triggeringrejection of the specimen.

Before or after the levels of the normalizing analytes are determined(but preferably after), the solution can be split into four aliquots, or“channels.” Each channel is then respectively tested for triglyceridelevel, HDL level, LDL level, and in a preferred embodiment, ALT level(which may be of interest in informing a physician whether the patienthas an abnormal liver which would contraindicate the use of certaindrugs). The analyte levels are measured using any technique known in theart or otherwise found to be suitable. For instance, a cholesterol testis disclosed in Allain, C. C., Poon, L. S., Chan, G. S. G., Richmond,W., and Fu, P. C., Clin. Chem. 20:474-75 (1974); see also Roeschlau, P.Brent, E. and Gruber, W A., Clin. Chem. Clin. Biochem. 12:226 (1974). Atest for HDL is disclosed in RiFai, N., Warnick, G. R., Ed., LaboratoryMeasurement of Lipids, Lipoproteins, and Apolioproteins (1994). A testfor triglycerides is disclosed in McGowan, M. W., Artiss, J. D.,Strandbergh, D. R., Zak, B. Clin. Chem. 29:583 (1983). A test for theliver enzyme ALT is disclosed in Wroblewski, F., LaDue, J. S., Proc.Sec. Exp. Biol. Med. 34:381 (1956). The invention is not limited to theforegoing tests or analytes, but to the contrary is applicable to othertests for these or other analytes.

After the analyte levels have been measured, the level of at least oneanalyte (and preferably all analytes) in the blood from which the bloodfluid specimen was obtained is calculated or otherwise determined basedon the solution level of the analyte and on the solution level of atleast one normalizing analyte. It is contemplated that the calculationof a blood analyte level may be as simple as multiplying the solutionanalyte level by the ratio of the blood normalizing analyte level to thesolution normalizing analyte level, the blood normalizing analyte levelbeing estimated based on the mean of a normal population distribution.For instance, it is believed that the normal blood sodium level inhumans ranges from 136 to 142 mEq/L with a mean of 139 mEq/L and thenormal chloride level ranges from 95 to 103 mEq/L with a mean of 99mEq/L. It is contemplated that through the use of two normalizinganalytes, the blood analyte level may be determined by calculating theblood analyte level based on the first normalizing analyte level,calculating the blood analyte level based on the second normalizinganalyte level, and then calculating the mean average of the bloodanalyte levels thus determined.

If additional normalizing analytes are evaluated, the mean average ofall blood level analytes thus determined may be calculated; if desired,where there are at least two normalizing analytes, the average may beweighted towards a specific normalizing analyte. For instance, it iscontemplated that Bayesian statistical methods may be used to assign arelative weight to the blood analyte levels determined with reference toeach analyte. Such statistical techniques may take into account not onlythe absolute magnitude of the level of the normalizing analyte level butalso the difference between the actual level and the magnitude expectedbased on the expected amount of serum, and the standard deviation of thenormal population distribution of the analyte. These techniques,sometimes referred to as “maximum likelihood” or “prior probabilityanalysis” techniques, may be used to provide an approximation of theblood analyte level. Further testing concerning such statisticaltechniques may be found in Casella, G., Berger, R. L., StatisticalInference (1990) and Carlin, B. P., Louis, T. A., Bayes and EmpiricalBayes Methods for Data Analysis (2d Ed. 2000).

Further details concerning the distribution of sodium, chloride, andosmolality in the normal human population may be found in Ravel,Clinical Laboratory Medicine (6th Ed. 1995); see also Penney, M. D. andWalters, G., Ann. Clin. Biochem. 24:566-71 (1987) and Fraser, C. G.,Cummings, S. T. Wilkinsen, S. O. et al., Clin Chem. 35: 783-86 (1985).It is further contemplated that a more complicated function of solutionanalyte level and the levels of one or more normalizing analytes may beemployed to calculate the blood analyte levels.

With reference now to FIG. 1, the generalized method shown therein isapplicable where the same entity performs the test and calculates theblood analyte level. Thus in steps 101 and 102 respectively the ISE(e.g., sodium) is immersed into the solution, and sodium levelinformation is obtained. The steps are repeated for the receipt ofchloride information, as shown in steps 103 and 104. Informationconcerning the analyte of interest is received in step 105, and theblood analyte level is calculated in step 106. If, in step 107, it isdesired to test an additional analyte for the same specimen, controlpasses to step 105 where the solution analyte information is receivedfor the new analyte. It is contemplated that the steps of testing forthe analytes of interest and the normalizing analytes may be performedby one entity and that the calculation of the blood analyte level may beperformed by a separate entity. Thus, for instance, in FIG. 1, steps 101and 103 may be omitted if the entity calculating the blood analyte levelis not the same entity as the entity that performs the test. The methodoutlined in FIG. 1 is very general, and other steps may be added, stepsmay be omitted or performed in a different order, and more generally themethod may be otherwise performed. For instance, steps of elution andverifying proper serum level are not shown, but are preferably employed.

It is contemplated that the analyte level, first normalizing analytelevel, and second normalizing analyte level may be independentlydetermined and these values used to calculate the blood level of theanalyte. For instance, the cholesterol tests hereinbefore discussedtypically are performed via enzymatic techniques in which the opticaldensity of a solution is measured. The “cholesterol value” of thesolution then may be expressed as:CV_(s) =f(OD)wherein CV_(s), the solution cholesterol concentration, is calculated asa function of the optical density, OD, when analytical reagents areadded to the sample in accordance with testing techniques known orotherwise found to be suitable. The solution sodium concentration, orNa_(s), may be used to calculate the blood cholesterol level, CV_(b), inthe following manner:CV_(b) =f(CV_(s),Na_(s))Numerous other forms of such calculations are possible. For instance, acorrection factor (CF) may be determined as a function of the solution'ssodium level, wherein:CV_(b) =f(CV_(s),CF)andCF=f(Na_(s))

It is alternatively contemplated that a single apparatus or system maybe designed for the calculation of blood analyte levels, wherein ananalog or digital electrical signal is generated corresponding to thelevels of analyte and normalizing analyte in the solution. For instance,the blood cholesterol number may be calculated as a function of themagnitude of two electrical signals:CV_(b) =f(E ₁ ,E ₂)wherein E₁ represents the magnitude of an electrical signal receivedfrom a spectrophotometer in measuring optical density for purposes ofevaluating total solution cholesterol level and E₂ represents themagnitude of an electrical signal received from an electrode specific tosodium.

In actual practice, it is contemplated that numerous variables willaffect the results obtained for a given set of specimens. For instance,the readings obtained from an ISE may “wander” from day to day, and thedevice used to collect the blood or other fluid specimen may containimpurities (such as sodium) that have the potential to introduce errorsinto the test. For this reason, from time to time a “tare” procedure maybe employed. Periodically, a plurality of specimens having a known ormeasurable analyte level is provided, and from these specimens areprepared modified specimens, the modified specimens being specimens asmodified in the manner expected of the unknown specimens. For instance,some number (e.g., six) blood specimens may be periodically placed ontoa blood spot collection device similar to those used in the field anddried, followed by elution of the dried samples to form solutions. Thesolutions are then tested for the level of the analyte and one or morenormalizing analytes. From these tests, an algorithm for determining theoriginal fluid analyte level as a function of the measured analyte leveland the levels of the normalizing analyte or analytes may be derived.Using this algorithm, modified fluid specimens may be analyzed, whereinthe levels of analyte and normalizing analyte may be measured, and thelevel of analyte in the original specimen may be determined as afunction thereof. Errors introduced by impurities (such as sodium) inthe collection device will be resolved by this methodology, and errorsintroduced by factors such as machine calibration will be resolvablewith periodic re-calculation of the algorithm. The tare procedure may beperformed occasionally or regularly at predetermined intervals (e.g.,every day, week, month, or year).

The foregoing exemplary equations are not meant to be exhaustive but, tothe contrary, are intended to illustrate that innumerable variants ofthe methods for calculating the blood analyte level are included withinthe scope of the invention. For instance, with respect to FIG. 2, in onesuch variant, an ISE (sodium) is immersed into an eluted sample at step201, and a signal corresponding to the sodium level is received at step202. The signal may be a digital signal, or may be an analog signal, thelevel of which is recorded. At steps 203 and 204, the same steps arerepeated for chloride level, and at steps 205 and 206 respectively, atest for the analyte is performed and a signal is received correspondingto the analyte level. At step 207, the solution sodium level iscalculated; at step 208, the chloride level is calculated, and at step209, the solution analyte level is calculated. At step 210, the bloodanalyte level is calculated, in this instance based on the magnitude ofthe solution sodium level, the solution chloride level, and the solutionanalyte level. If, at step 211, it is desired to test for an additionalanalyte for the same specimen, control passes to step 205. In such case,if the solution sodium and chloride level have been stored, steps 207and 208 may be omitted after a signal is received corresponding to thesecond analyte level. The process may be controlled by any suitablemicroprocessor or microcontroller (not shown).

As stated hereinabove, it is contemplated that the entity who providestest results to a user, who may or may not be the health careprofessional who has ordered the test, in turn may be the same ordifferent entity from the entity which performs the calculation of theblood analyte level, which in turn may be the same or different entityfrom the entity which tests the specimen and generates informationcorresponding to the analyte level or levels and the normalizing analytelevel or levels. A very general protocol for a results providingfacility is set forth in FIG. 3, wherein an inquiry is received from auser at step 301, and the user is prompted for his or her test number atstep 302. At step 303, the test number is received, and at step 304, atest result database is queried for test result information. Theinformation is received at step 305 and is provided to the user at step306.

With further reference to FIG. 4, the test result database describedabove may be structured in any suitable manner. With respect to, forinstance, database record 400, the test result information 401, which inthe illustrated embodiment includes two items of information, bloodanalyte information 1 and blood analyte information 2, is correlatedwith the test number 402. The test number may be an anonymous testnumber or may be a test number that is associated with a user, forinstance, elsewhere in the database record 400 (not shown) or in adifferent database.

With reference to FIG. 5, the database may be prepared by creating adatabase record (shown in step 501), receiving test result informationand a test number (shown in steps 502 and 503 respectively) and, asshown in step 504, entering the test number and test result informationinto the database record. More information concerning the role of aresults providing facility in a medical or wellness testing protocol canbe found in the aforementioned Quattrocchi patents and in copendingapplication Ser. No. 09/709,884.

The invention additionally contemplates a method for preparing adatabase for use in calculating blood analyte levels. The blood analytelevel may be calculated with specific reference to the database, oralternatively the database may be used in conjunction with thepreparation of an algorithm for enabling blood level calculation. Thedatabase preferably is prepared with reference to blood having a knownlevel of cholesterol or other analyte of interest. Plural specimens ofblood having different levels of the analyte are then reduced to anmodified specimen, such as a blood spot or serum specimen, and eachspecimen is analyzed for the analyte of interest and for a normalizinganalyte. For instance, with respect to FIG. 6, a database record iscreated at step 601, and known blood analyte level information isreceived at step 602. Information as to the solution analyte level andthe level of two normalizing analytes, sodium and chloride, for example,are received at steps 603-605, and at step 606, the information receivedis entered into the database record. If, at step 607, an additionaldatabase record is to be created, control passes to step 601, wherein anew database record is created for the new specimen. It should be notedthat the order of the steps is not critical, and indeed the database maybe prepared sequentially with respect to each blood specimen (i.e., eachspecimen is reduced to an modified specimen, tested, and the resultsentered into a database record prior to altering the next specimen ofblood), sequentially with respect to database record (wherein all of theblood specimens are reduced to modified specimens prior to entering thefirst database records) or by any other suitable methodology. A databaserecord 700 as shown in FIG. 7 is thus prepared, with entries 701 through704 representing respectively blood analyte level, solution analyte,solution sodium level, and solution chloride level.

As discussed above, rather than being calculated, the blood analytelevel in a blood fluid specimen may be determined with reference to thedatabase, for instance, by finding the solution analyte level andsolution normalizing analyte level or levels in the database that areclosest to those of the specimen. Alternatively, any suitablestatistical or mathematical technique may be used to derive an algorithmfor calculating the blood analyte level from the solution analyte leveland at least one normalizing analyte level. In some embodiments, thealgorithm is first order with respect at least to the solution analytelevel, and may be first order with respect to the solution analyte leveland one or both normalizing analyte levels.

The invention preferably is conducted in accordance with the generalschematic set forth in FIG. 8. Generally the customer 801 purchases atest kit from a physician or retail store 802 (transfer of the kit isshown via transfer communication 805). The test kit (not shown)preferably includes instrumentalities for allowing the customer toobtain a blood, serum or serum spot specimen. For instance, as discussedmore fully in the aforementioned Quattrocchi patents, the test kit mayinclude a lancet for pricking the user's finger, a blood spot card, orserum spot card, (or the device shown in FIGS. 9 and 10 hereinafterdiscussed) an informed consent form, and a test number. After preparingthe blood, serum or serum spot card, the customer sends the dried bloodspecimen to a results providing facility 803 as shown via transfercommunication 806. In the illustrated embodiment, the results providingfacility 803 sends the specimen to a separate testing facility 804, asshown via transfer communication 809. As shown via communication 810,the testing facility provides the test results to the results providingfacility. The results may be “raw” results, i.e., results in which thelevel of the analyte in the blood has not been determined or obtained,or alternatively the testing facility may calculate the blood analytelevel and report that result to the results providing facility. As shownat communication 807, the customer contacts the results providingfacility, and at communication 808, the results providing facilityprovides the test results to the customer. Optionally, the resultsproviding facility may be equipped to communicate directly with thephysician's office, as shown at communications 811 and 812. Except wheretransfer of a physical specimen is required, the communication may bemade via any means or method now known or hereinafter discovered, forinstance, via telephone, wireless communication, electronic mail or“chat” or other electronic communication, or other form ofcommunication.

With reference now to FIGS. 9 and 10, the illustrated fluid collectiondevice 900 includes two gangs 901, 901, each comprising a fluidcollector 903, 904 that is disposed between a superstrate sheet 905 anda substrate sheet 906 and that is generally fixed with respect to thesuperstrate sheet 905. The fluid collector is ordinarily connected tothe substrate sheet 906 (a portion of which is visible) at one end 907,908, although the collector may be flexible and thus not entirely fixedwith respect to the substrate sheet 905. The substrate is provided withat least one aperture (two shown as 909, 910) by which a user mayfluidically transfer blood to the collector. In the illustratedembodiments, secondary apertures 911, 912 are provided. To use thedevice, a user dispenses blood onto the collector, whereby some or allof the blood wicks in the direction shown by arrow 913 until theportions 914, 915 of the collectors 903, 904 visible through thesecondary apertures 914, 915 become tainted, whereupon the user isprovided with an indication that sufficient blood has been collected. Inthe illustrated embodiments, instructions 917 are provided on thesubstrate sheet 905 and identification information spaces 918 (shown inFIG. 10) are provided on the substrate sheet 906. The device may beprovided with non-textual machine-readable indicia (such as barcode919).

In a highly preferred embodiment of the invention, the fluid collectoris an absorbent paper or glass fiber substrate that is coated with asaccharide, preferable a mono- or di-saccharide and most preferablyxylose. The substrate should be one that permits at least substantialseparation of the red blood cell component of blood cells from otherportions of the blood (i.e., serum). It is believed that the saccharidecomponent permits more effective recovery of the serum components fromthe substrate sheet. The substrate may be coated only at the surface onone or both sides with the saccharide, but preferably the substrate iscoated on internal surfaces as well as on the exterior surface. In oneembodiment, 180 μl of a 5% solution of xylose is applied to the internalsurface of the 0.8×7 cm substrate (such that substantially all of thesubstrate is wetted) and allowed to air dry. If the fluid collector isused in the device shown in FIGS. 9 and 10, the blood cells will remainnear the end of the collection device (opposite the direction of arrow913) while the serum will wick toward the other end of the card. Uponreceipt by a testing laboratory, a portion of the fluid collector may beexcised and eluted. Preferably, the excised portion includes a portionof the collector “above” the terminal wicking point of the serum. Onecommercial product (Whatman GF/AVA paper) contains sodium, and it isbelieved that by excising filter paper above the terminal wicking pointa consistent amount of sodium will be introduced into the eluted fluid.The device may be prepared by applying a solution of the saccharide tothe substrate

The invention enables venous blood analyte levels to be determined fromcapillary blood specimens. It is contemplated that in most embodimentsthe solution analyte level will be normalized to the venous blood levelof the analyte, but it is also contemplated that the solution value maybe normalized to capillary blood level (or for that matter a differentblood level).

The databases discussed herein may be created and stored as computerfiles on a computer readable medium, such as a diskette, hard disk,CD-ROM, DVD-ROM, ROM chip or EPROM chip, or any other suitable medium asmay be now known or hereinafter discovered. The tests for the analyteand normalizing analytes may be performed by any conventional orotherwise suitable technique now or hereinafter found to be suitable,and likewise the analyte and normalizing analyte (which may be discreteatoms, ions, compounds, biochemical materials, or properties) may bethose specifically described herein or others as may be found suitablefor use in conjunction with the invention.

The following examples are provided to illustrate the invention, butshould not be construed as limiting the invention in scope unlessotherwise indicated. Unless otherwise indicated in these examples, themeasured analyte level was corrected using sodium as the solenormalizing analyte. The correction was made using a simple linearregression. It should be understood that more complex single variableand multivariate regressions may be used in conjunction with theinvention, and thus the statistical techniques employed in theseexamples should be viewed as non-limiting.

EXAMPLE 1

This example demonstrates the performance of the invention in themeasurement of total cholesterol.

Fifteen patients were used to obtain blood specimens (micro-serumspecimens) via venal puncture. Serum from each specimen was spotted anddried on filter paper with applied volumes ranging from approximately 8to 16 μl. The number of spots for each blood specimen is listed in thecolumn “No.” in the table below. Each spot was eluted and measured forcholesterol and sodium. For each specimen for each patient, thenormalized cholesterol level was calculated based on the level of ameasured analyte in the fluid (cholesterol) and a normalizing analyte(sodium). The normalized cholesterol level was obtained according to thepresent invention using linear regression techniques to yield thefollowing function: Normalized Cholesterol=MeasuredCholesterol/((−0.003306)+0.9781×(Measured Sodium/139)), where 139(mEq/L) is the population mean for sodium. The regression was calculatedbased on five direct measurements of the cholesterol level from the sameblood sample, as listed in the column “Mean Serum Cholesterol.” The meanaverage of the normalized cholesterol values for each patient is givenin the column “mean normalized cholesterol” and the coefficient ofvariation of the normalized cholesterol levels obtained for each patientis listed in the column designated “Normalized Cholesterol CV %.”

Normalized Mean Serum Mean Normalized Cholesterol Patient No.Cholesterol Cholesterol CV % A 11 152.35 153.68 3.85 Ja 12 165.79 162.501.42 Il 14 180.93 180.47 4.61 Ca 12 186.20 182.28 0.70 Br 10 187.06185.35 2.93 Mi 12 187.14 186.21 1.85 Gr 12 187.42 189.14 1.65 Ed 12200.38 197.18 1.36 Tr 11 220.83 221.89 2.00 Bb 11 232.65 233.06 1.89 Ma11 236.73 245.53 1.02 Jo 11 237.37 237.24 1.95 JJ 14 262.41 259.24 1.75Kt 12 264.30 268.23 1.86 TT 13 269.36 273.53 2.79

A comparative linear regression was generated for the data pointscollected in this Example. The linear fit followed the followingequation:Mean Normalized Cholesterol=−7.97+1.04×Mean Serum Cholesterol,with the correlation coefficient, expressed as R², being greater than0.99.

EXAMPLE 2

This example demonstrates the performance of the invention in themeasurement of HDL.

The same dried spots from the same fifteen patients in Example 1 wereused to obtain a measured value for HDL. The normalized HDL level wasobtained according to the present invention using linear regressiontechniques yielding the following function: NormalizedHDL=HDL/(0.0158+1.060×(Sodium/139)). The following data was measured orcalculated in the same manner as in Example 1.

Mean Normalized Normalized Patient No. Mean Serum HDL HDL HDL CV % II 1445.77 47.03 2.35 A 11 46.05 47.77 2.17 Jo 11 47.40 48.50 2.12 Ja 1248.87 53.22 2.23 JJ 14 49.07 48.15 1.68 Gr 12 49.64 52.45 1.62 Mi 1259.96 58.95 1.69 Br 10 57.20 55.83 2.66 Ed 12 71.00 71.09 0.92 Kt 1273.08 72.46 1.53 TT 13 76.16 75.77 2.27 Ca 12 78.01 75.93 1.50 Bb 1178.77 73.35 1.99 Ma 11 87.84 84.75 0.94 Tr 11 91.15 86.42 1.46

A comparative linear regression was generated for the data pointscollected in this Example. The linear fit followed the followingequation:Mean Normalized HDL=8.15+0.87×Mean Serum HDL,with the correlation coefficient, expressed as R², being greater than0.99.

EXAMPLE 3

This example demonstrates the performance of the invention in themeasurement of triglycerides (TG).

The same dried spots from the same fifteen patients in Example 1 wereused to obtain a measured value for TG. The normalized TG level wasobtained according to the present invention using linear regressiontechniques yielding the following function: NormalizedTG=TG/((−0.0136)+0.9307×(Sodium/139)). The following data was measuredor calculated in the same manner as in Example 1.

Mean Normalized Normalized Patient No. Mean Serum TG TG TG CV % Ca 1237.63 38.76 1.95 Bb 11 46.86 48.55 1.75 A 11 48.75 50.16 2.73 Ja 1249.68 49.94 3.31 Kt 12 52.15 48.19 1.32 Br 10 55.00 56.56 4.14 Ma 1156.05 56.40 2.03 II 14 59.09 60.88 6.22 Ed 12 62.91 61.65 1.25 Tr 1166.69 67.66 1.63 TT 13 68.76 72.14 13.37 Mi 12 71.84 72.63 1.62 Jo 11109.28 107.10 2.27 JJ 14 117.31 112.24 5.03 Gr 12 139.47 136.74 2.13

A comparative linear regression was generated for the data pointscollected in this Example. The linear fit followed the followingequation:Mean Normalized TG=3.36+0.95×Mean Serum TG,with the coefficient, expressed as R², being greater than 0.995.

EXAMPLE 4

This example demonstrates the performance of the invention in themeasurement of LDL. The same observations from the same fifteen patientsin Example 1, 2 and 3 were used to calculate a value for LDL in serumand a value for LDL in MSS according to the Friedewald formula:Mean Serum LDL=Mean Serum Cholesterol−Mean Serum HDL−Mean Serum TG/5Mean Normalized LDL=Mean Normalized Cholesterol−Mean Normalized HDL−MeanNormalized TG/5, respectively.The following data was calculated (mean serum LDL was calculated fromthe mean values reported in Examples 1-3)

Mean Normalized Normalized Patient No. Mean Serum LDL LDL LDL CV % A 1196.55 95.30 5.36 Ca 12 100.66 98.82 1.17 Ja 12 106.98 99.22 2.32 Gr 12109.88 109.00 2.19 Mi 12 115.81 112.90 2.80 Tr 11 116.35 121.21 3.11 Ed12 116.80 113.76 1.98 Br 10 118.86 118.21 3.23 Il 14 123.34 119.75 5.72Ma 11 137.68 149.45 1.72 Bb 11 144.51 150.01 2.12 Jo 11 168.11 167.552.66 Tt 13 179.45 183.33 3.33 Kt 12 180.78 186.13 2.19 JJ 14 189.88189.54 1.89

A comparative linear regression was generated for the data pointscollected in this Example. The linear fit followed the followingequation:Mean Normalized LDL=−8.16+1.07×Mean Serum LDL,with the correlation, expressed as R², being equal to 0.98.

EXAMPLE 5

This example demonstrates the performance of the invention in themeasurement of total cholesterol.

One hundred thirty-two patients were used to obtain blood via venalpuncture (venous blood specimens) and by pricking their fingers(capillary blood specimens). Capillary blood was spotted onxylose-coated Whatman GF/AVA filter paper, using a device similar tothat shown in FIG. 9. Capillary blood specimens were dried and theportion of the filter paper which contained separated serum was cut outand eluted. Eluate from each specimen was measured for cholesterol andsodium. The normalized cholesterol level was obtained according to thepresent invention using a variable formula: NormalizedCholesterol=Measured Cholesterol/(A+B×(Measured Sodium/139)). In thisequation, A and B were scalar values that were periodically recalculatedbased on the “tare” procedure heretofore described, whereby a regressionfor six patients was calculated and the A and B values from thisregression were used to calculate normalized cholesterol values forspecimens analyzed before the next tare period. Actual (directlymeasured in venous blood) and calculated normalized cholesterol valvesfor these patients are given below.

Serum Normalized Patient Cholesterol Cholesterol 1 172.68 157.54 2149.25 154.61 3 176.81 175.60 4 189.78 187.41 5 170.38 173.03 6 189.67188.80 7 130.52 128.80 8 266.76 276.31 9 151.29 152.49 10 219.86 211.2311 242.00 251.07 12 232.41 230.66 13 173.09 176.48 14 190.89 190.86 15264.47 260.46 16 236.18 244.49 17 272.58 279.76 18 240.29 228.83 19169.32 166.57 20 192.02 195.03 21 239.83 235.33 22 225.13 225.13 23169.40 156.05 24 197.93 183.67 25 151.59 146.26 26 235.43 247.88 27178.84 170.79 28 196.40 191.34 29 240.99 230.52 30 171.53 173.95 31229.43 229.43 32 217.54 223.84 33 187.23 183.58 34 175.68 173.95 35174.69 172.34 36 251.23 249.20 37 203.70 185.98 38 123.30 114.96 39136.04 127.97 40 251.33 243.27 41 216.14 218.02 42 145.14 156.86 43208.58 203.43 44 250.25 245.07 45 235.76 250.40 46 193.19 187.83 47211.75 223.38 48 221.15 226.04 49 199.41 196.35 50 249.35 259.44 51166.46 165.63 52 154.64 151.56 53 187.36 190.37 54 256.78 260.40 55230.59 222.39 56 208.57 224.14 57 183.92 181.28 58 159.73 156.20 59155.31 153.59 60 205.29 197.61 61 204.49 198.97 62 219.21 221.45 63122.83 114.88 64 175.13 176.48 65 201.35 211.70 66 216.66 209.09 67227.50 231.96 68 151.28 153.23 69 130.10 128.40 70 175.95 173.45 71182.38 183.21 72 201.03 195.89 73 175.86 189.73 74 146.10 149.88 75116.17 103.88 76 193.58 197.59 77 291.91 296.11 78 184.93 185.49 79145.82 141.34 80 182.73 180.78 81 175.84 170.03 82 148.99 151.67 83212.79 213.40 84 228.82 225.39 85 218.44 229.26 86 169.43 173.84 87151.43 157.96 88 217.96 218.63 89 239.39 244.17 90 148.62 152.86 91136.81 132.60 92 179.13 173.31 93 121.10 119.61 94 165.37 163.34 95117.65 132.34 96 190.25 184.44 97 201.78 206.49 98 133.26 137.69 99225.84 221.67 100 244.66 230.25 101 164.72 168.10 102 150.75 146.82 103163.57 170.41 104 196.06 198.89 105 213.32 206.01 106 186.62 183.13 107163.46 162.71 108 244.58 250.24 109 231.82 231.32 110 171.94 172.27 111207.12 209.36 112 205.41 209.00 113 157.54 156.02 114 191.41 190.59 115192.20 197.37 116 193.52 183.72 117 257.83 248.49 118 178.32 171.44 119203.64 209.32 120 210.36 230.25 121 207.74 220.04 122 200.05 205.38 123216.34 219.09 124 190.10 179.14 125 293.34 272.48 126 228.57 226.02 127171.60 174.88 128 142.80 148.94 129 197.16 205.05 130 220.50 218.43 131220.32 231.50 132 255.78 255.23

A comparative linear regression was generated for the data pointscollected in this Example. The linear fit followed the followingequation:Normalized Cholesterol=−1.16+1.00×Serum Cholesterol, with thecorrelation coefficient, expressed as R ², being 0.966.

EXAMPLE 6

This example demonstrates the performance of the invention in themeasurement of HDL. The dried spots and venous blood specimens from thesame one hundred thirty-two patients in Example 5 were used to measureHDL in capillary blood and compare it to a measured value for HDL invenous blood. The normalized HDL level in capillary blood was obtainedaccording to the present invention using a formula: NormalizedHDL=Measured HDL/(A+B×(Measured Sodium/139)), where A and B wereobtained as previously described. The following results were observed.

Serum Normalized Patient HDL HDL 1 58.90 61.12 2 41.28 42.33 3 38.5439.15 4 48.84 46.19 5 61.56 54.98 6 52.68 48.79 7 47.69 45.15 8 34.6939.49 9 57.45 56.32 10 38.00 36.33 11 47.53 42.14 12 60.04 58.94 1336.08 37.35 14 46.09 48.37 15 42.22 42.82 16 34.70 38.98 17 55.76 55.7918 21.16 24.53 19 55.33 55.69 20 44.66 42.65 21 83.26 81.00 22 44.3346.14 23 40.71 40.69 24 47.24 43.98 25 49.46 47.71 26 44.37 43.30 2750.16 48.34 28 55.49 61.30 29 58.90 61.12 30 41.28 42.33 31 38.54 39.1532 48.84 46.19 33 61.56 54.98 34 52.68 48.79 35 47.69 45.15 36 34.6939.49 37 57.45 56.32 38 38.00 36.33 39 47.53 42.14 40 60.04 58.94 4136.08 37.35 42 46.09 48.37 43 42.22 42.82 44 34.70 38.98 45 55.76 55.7946 21.16 24.53 47 55.33 55.69 48 44.66 42.65 49 83.26 81.00 50 44.3346.14 51 40.71 40.69 52 47.24 43.98 53 49.46 47.71 54 44.37 43.30 5550.16 48.34 56 55.49 61.30 57 49.27 45.94 58 51.73 51.78 59 38.07 36.9860 38.22 38.49 61 43.57 45.05 62 54.16 51.46 63 38.66 34.13 64 50.1448.65 65 57.94 54.11 66 46.02 44.67 67 49.21 52.36 68 43.15 45.31 6937.20 38.42 70 49.66 50.00 71 63.00 65.28 72 79.92 79.17 73 37.12 44.5774 59.14 60.35 75 32.49 28.57 76 56.08 59.37 77 64.22 70.04 78 46.5448.66 79 37.68 37.28 80 75.41 74.70 81 44.06 44.73 82 40.65 40.88 8393.40 91.97 84 40.97 47.04 85 69.63 75.17 86 36.13 38.81 87 34.88 36.4288 43.90 49.40 89 63.29 66.41 90 49.21 49.65 91 29.54 31.27 92 49.3049.87 93 35.82 34.39 94 49.66 51.20 95 39.01 39.79 96 36.92 34.49 9743.40 43.45 98 48.70 45.97 99 42.15 41.04 100 59.09 55.11 101 49.4647.04 102 33.36 29.81 103 49.36 47.93 104 43.02 39.12 105 39.81 41.06106 60.29 56.62 107 59.84 55.33 108 84.77 82.31 109 55.20 55.72 11054.77 56.06 111 69.16 67.30 112 38.18 40.50 113 37.11 36.49 114 51.3149.24 115 39.69 42.54 116 61.17 56.56 117 29.94 30.25 118 75.50 77.62119 56.94 57.49 120 68.89 71.30 121 37.89 40.82 122 73.57 72.14 12378.31 78.16 124 48.88 47.45 125 83.96 79.26 126 95.12 92.48 127 51.4452.50 128 38.88 38.10 129 41.70 44.58 130 47.80 46.24 131 56.42 59.35132 55.14 56.98

A comparative linear regression was generated for the data pointscollected in this Example. The linear fit followed the followingequation:Normalized HDL=2.47+0.953×Serum HDL,with the correlation coefficient, expressed as R², being greater than0.96.

EXAMPLE 7

This example demonstrates the performance of the invention in themeasurement of triglycerides (TG). The dried spots and venous bloodspecimens from the same one hundred thirty-two patients in Example 5were used to measure TG in capillary blood and compare it to a measuredvalue for TG in venous blood. The normalized TG level in capillary bloodwas obtained according to the present invention using the formula:Normalized TG=Measured TG/(A+B×(Measured Sodium/139)), where A and Bwere obtained as previously described. The following results wereobserved.

Serum Normalized Patient TG TG 1 73.24 55.65 2 97.89 97.31 3 45.26 38.384 70.31 60.30 5 119.71 119.33 6 105.97 100.56 7 77.47 73.30 8 220.18236.94 9 191.79 203.18 10 177.10 177.03 11 112.19 116.71 12 73.24 55.6513 97.89 97.31 14 45.26 38.38 15 70.31 60.30 16 119.71 119.33 17 157.70164.69 18 122.09 124.56 19 66.86 63.24 20 138.31 151.08 21 146.08 137.3622 95.85 97.05 23 77.27 60.69 24 85.44 82.87 25 86.25 77.32 26 112.51110.68 27 176.25 184.16 28 190.63 189.57 29 95.17 98.92 30 98.52 98.7631 102.13 97.07 32 117.77 128.91 33 123.08 125.56 34 135.72 132.69 3576.46 71.14 36 230.90 210.77 37 80.41 67.66 38 99.43 85.63 39 86.8791.07 40 125.01 120.98 41 362.90 322.04 42 132.98 118.47 43 83.21 75.4344 52.45 53.34 45 53.91 50.52 46 349.76 357.87 47 135.25 139.57 48209.20 208.33 49 374.36 386.86 50 74.90 79.81 51 395.31 399.34 52 56.3854.87 53 217.08 258.78 54 52.83 71.35 55 136.53 144.81 56 115.98 118.4557 78.41 62.45 58 70.38 65.13 59 91.00 68.59 60 180.98 179.72 61 163.32188.88 62 72.16 65.05 63 102.89 101.45 64 50.24 49.05 65 184.45 195.4266 183.07 194.25 67 65.28 65.04 68 111.40 109.43 69 67.25 87.27 70 74.9272.25 71 100.19 105.33 72 136.82 132.52 73 119.29 129.90 74 119.76119.83 75 121.90 125.90 76 75.55 80.65 77 74.44 89.06 78 226.78 243.0579 71.19 78.23 80 98.89 93.66 81 127.93 135.56 82 333.65 352.31 83 97.1891.96 84 139.77 133.20 85 73.23 72.05 86 160.00 148.64 87 131.69 133.4988 69.07 66.79 89 271.22 248.43 90 91.86 98.00 91 231.14 224.76 92153.65 171.85 93 115.95 107.16 94 263.50 257.68 95 95.38 92.85 96 143.96125.21 97 110.10 131.36 98 97.72 93.75 99 158.22 151.23 100 123.80127.26 101 279.56 271.61 102 192.26 176.02 103 59.41 59.23 104 197.04186.32 105 182.29 170.98 106 96.16 91.53 107 80.46 72.56 108 65.55 68.16109 215.37 210.92 110 186.09 191.14 111 96.41 96.52 112 78.68 80.54 11383.96 73.13 114 207.32 208.03 115 37.41 37.32 116 103.17 93.38 117193.21 210.21 118 119.46 103.27 119 67.57 58.99 120 119.56 117.34 12175.42 52.90 122 311.18 315.01 123 67.72 68.28 124 127.36 129.28 12559.82 64.57 126 85.54 83.90 127 43.24 41.49 128 85.09 78.05 129 95.1599.45 130 92.21 75.05 131 72.46 88.51 132 56.52 57.13

A comparative linear regression was generated for the data pointscollected in this Example. The linear fit followed the followingequation:Normalized TG=−2.5+1.01×Serum TG,with the coefficient, expressed as R², being 0.98.

EXAMPLE 8

This example demonstrates the performance of the invention in themeasurement of LDL. The same observations from the same one hundredthirty-two patients in Example 5, 6 and 7 were used to calculate a valuefor LDL in serum and a value for LDL in MSS according to the Friedewaldformula:Serum LDL=Serum Cholesterol−Serum HDL−Serum TG/5Normalized LDL=Normalized Cholesterol−Normalized HDL−Normalized TG/5.The following results were calculated:

Serum Normalized Patient LDL LDL 1 110.85 101.31 2 97.93 101.60 3 109.82103.89 4 110.51 112.89 5 108.21 107.74 6 126.07 121.49 7 49.76 54.13 8173.19 173.53 9 78.72 77.71 10 129.52 119.61 11 174.74 180.58 12 149.04140.09 13 98.72 101.13 14 115.22 115.25 15 187.28 180.56 16 146.29158.21 17 195.20 200.00 18 167.30 161.22 19 101.87 99.92 20 122.26127.45 21 168.27 164.79 22 149.27 148.28 23 100.92 85.06 24 129.40115.93 25 92.09 88.71 26 165.02 175.92 27 114.88 104.61 28 94.16 90.6229 154.94 142.87 30 114.95 117.39 31 144.71 148.13 32 152.62 164.12 33105.78 111.48 34 105.62 106.95 35 101.99 102.99 36 143.96 145.31 37119.65 105.96 38 68.65 63.55 39 78.02 75.16 40 180.50 174.23 41 110.10109.10 42 72.00 80.58 43 124.52 118.94 44 140.68 128.72 45 165.02 178.6646 92.96 83.43 47 145.15 156.72 48 133.07 131.63 49 105.59 101.08 50177.71 184.34 51 102.56 101.25 52 91.72 95.09 53 123.82 129.64 54 194.21203.38 55 144.23 138.11 56 120.42 125.06 57 120.22 122.32 58 87.42 84.1359 107.19 106.80 60 130.18 120.03 61 124.30 115.07 62 151.99 156.98 6361.89 58.86 64 111.54 110.38 65 128.43 143.14 66 150.60 143.35 67 150.93153.10 68 84.28 81.94 69 68.95 66.01 70 101.92 98.27 71 104.27 101.79 72106.22 98.91 73 93.38 96.56 74 72.72 73.88 75 63.90 56.58 76 111.92111.11 77 160.96 155.60 78 118.95 118.43 79 80.19 77.42 80 92.68 91.6781 99.78 95.56 82 82.00 84.09 83 105.58 108.07 84 133.61 128.66 85130.44 134.49 86 87.08 90.07 87 85.82 87.16 88 150.87 147.80 89 123.40126.22 90 80.33 84.64 91 78.47 76.28 92 107.81 97.16 93 65.74 66.47 9484.06 81.90 95 53.88 67.10 96 97.42 95.63 97 119.93 127.83 98 72.6979.87 99 144.28 143.36 100 149.12 140.94 101 96.03 102.76 102 101.29102.50 103 101.09 108.85 104 109.96 117.58 105 136.29 126.72 106 107.05107.20 107 87.89 91.28 108 143.01 153.30 109 135.15 134.00 110 109.68108.74 111 117.33 123.38 112 128.59 126.45 113 96.53 98.88 114 126.58129.56 115 128.60 131.36 116 117.26 116.58 117 165.65 155.24 118 89.2780.15 119 121.23 125.97 120 129.51 146.03 121 152.74 162.44 122 117.83124.94 123 121.01 125.32 124 122.19 111.80 125 190.94 178.21 126 118.95115.83 127 108.85 110.95 128 76.17 84.41 129 116.38 120.15 130 116.39113.82 131 134.85 142.75 132 173.09 172.40

A comparative linear regression was generated for the data pointscollected in this Example. The linear fit followed the followingequation:Normalized LDL=−0.25+1.00×Serum LDL,with the correlation, expressed as R², being equal to 0.96.

It is thus seen that the invention provides a method for determining thelevel of an analyte in a specimen.

While particular embodiments to the invention have been describedherein, the invention is not limited thereto, but to the contrary shouldbe deemed defined by the full scope of the appended claims. Allreferences and pending applications cited herein are hereby incorporatedby reference in their entireties.

1. A method for determining a level of an analyte from a solution formedfrom a dried blood fluid specimen eluted with a fluid, comprising: inany order, measuring a first level of an analyte in said solution,measuring the level of a first normalizing analyte in said solution, andmeasuring the level of a second normalizing analyte in said solution;and calculating the level of the analyte in the blood from which saidblood fluid specimen was collected based on said first level of theanalyte in said solution, the level of said first normalizing analyte insaid solution, and the level of said second normalizing analyte in saidsolution by calculating an average of the level of said first and secondnormalizing analyte in said solution.
 2. The method of claim 1, whereinsaid analyte is a cholesterol.
 3. The method of claim 1, wherein saidanalyte is a ALT.
 4. The method of claim 1, wherein said analyte is PSA.5. The method of claim 1, wherein said first or second normalizinganalyte is chloride.
 6. The method of claim 1, wherein said first orsecond normalizing analyte is osmolality.
 7. The method of claim 1,further comprising: calculating the first level of analyte in saidsolution; and calculating the level of the analyte in the blood fromwhich said blood fluid specimen was collected by multiplying said firstlevel of analyte in said solution by a correction factor, saidcorrection factor being calculated as a function of the level of one ofthe normalizing analytes in said solution.
 8. A method of reporting atest result comprising a level of analyte calculated according to themethod of claim 1, the method comprising: receiving an incoming inquiryfrom a user; prompting said user for a test number; retrieving a testresult from a database of test numbers and test results; and reportingsaid test results to said user.
 9. A method for preparing a database oftest results comprising a level of analyte calculated according to themethod of claim 1, the method comprising: receiving a test number;receiving a test result; and storing said test result and said testnumber in a database record.
 10. The method of claim 1, wherein saidfirst and second normalizing analytes are dissimilar and are eachselected from the group consisting of sodium, chloride, and osmolality.